Effect of rolipram and dibutyryl cyclic AMP on resequestration of cytosolic calcium in FMLP-activated human neutrophils

1 We have investigated the effects of the selective phosphodiesterase (PDE) type 4 inhibitor, rolipram (0.01-1 mu M) on cytosolic Ca2+ fluxes in FMLP-activated human neutrophils, as well as on superoxide production by, and release of elastase from, these cells. 2 Cytosolic Ca2+ fluxes were measured by use of fura-a spectrofluorimetry in combination with a radiometric procedure that enables distinction between net efflux and influx of the cation. Superoxide production and elastase release were measured by lucigenin-enhanced chemiluminescence and a colorimetric procedure, respectively. 3 Pretreatment of neutrophils with rolipram did not affect the FMLP-activated release of Ca2+ from intracellular stores, but was associated with dose-related acceleration of the rate of decline in fura-2 fluorescence and with decreased efflux, as well as store-operated influx of Ca-45(2+), indicative of enhancement of resequestration of the cation by the endo-membrane Ca2+-ATPase. 4 Inhibition of superoxide production and elastase release was observed at concentrations of rolipram which accelerated the clearance of Ca2+ from the cytosol of FMLP-activated neutrophils. 5 These effects of rolipram on FMLP-activated Ca2+ fluxes, superoxide generation and elastase release were mimicked by pretreatment of neutrophils with dibutyryl cyclic AMP (0.5-4 mM), while theophylline (10-150 mu M), a non-specific PDE inhibitor, as well as the beta(2)-agonist, salbutamol, were less effective. 6 We conclude that rolipram deactivates FMLP-stimulated human neutrophils by enhancement of cyclic AMP-dependent resequestration of cytosolic Ca2+.